Targeting BCR-ABL with specific kinase inhibitors has resulted in remarkable clinical advances and has dramatically changed the outcome of patients with CML and Ph+ ALL. Despite that, the emergence of leukemic cell resistance raises serious concerns and the need for the development of approaches to overcome this resistance. It is also important to note that there is now evidence that, TKIs do not eliminate leukemia initiating stem cells (LICs), even in sensitive cases where complete remission is achieved. Thus, efforts to target cellular pathways down-stream of BCR-ABL, circumventing resistance at the BCR-ABL level, may provide an important clinical approach for the treatment of such Ph+ leukemias and the elimination of LICs. Work from our laboratory has established that the mTOR pathway is deregulated in Ph+ leukemias and has raised the possibility that such deregulation contributes to the emergence of leukemic cell resistance. We have provided evidence on the existence of two functionally distinct mTORC1 complexes in BCR-ABL transformed cells, rapamycin-sensitive (RS) and rapamycin-insensitive (RI) mTORC1, and shown that RI-mTORC1 plays critical roles in the regulation of mRNA translation for oncogenic proteins that promote leukemic cell proliferation. We have also established that mTORC2 complexes are formed and their activation is important for survival of leukemia cells and primary Ph+ leukemic precursors. Using a unique dual catalytic inhibitor of mTORC1 and mTORC2, OSI-027, we have established that targeting such complexes results in potent suppressive effects on primitive leukemic precursors from CML patients and cells expressing the T315I BCR- ABL mutation. In parallel efforts to target the AMPK metabolic-sensor pathway in Ph+ cells, we found that AMPK inducers suppress RI-mTORC1 complexes, resulting in potent antileukemic effects. Such agents also overcome resistance in cells expressing refractory BCR-ABL mutations, such as T315I. These findings raise the prospect of future translational approaches to overcome resistance in CML and Ph+ ALL by directly targeting RI-mTORC1 and mTORC2 complexes and/or by engaging AMPK. The current proposal is a systematic approach to define the mechanisms of deregulation of mTOR pathways in Ph+ leukemias and to identify downstream effectors that could be therapeutically targeted. In addition it involves studies to identify feedback pathways that account for leukemic cell resistance and to use agents that target such pathways to eliminate LICs in Ph+ leukemias. Specific aim 1 will dissect BCR-ABL-regulated signaling events that control mTORC2 and RI-mTORC1 complexes and will define the regulatory effects of AMPK modulation in the process. Specific aim 2 will identify downstream effectors of mTORC1 and mTORC2 complexes and will systematically define the relevance of targeting distinct effectors in the generation of antileukemic responses. Specific aim 3 will employ CML mouse models for TKI sensitive and resistant (T315I- BCR-ABL) CML to examine the in vivo antileukemic properties of mTORC1/2 targeting agents. Finally, specific aim 4 will systematically study the effects of OSI-027 and AMPK activators on primary cells from a large number of patients with CML and Ph+ ALL and their effects on survival of leukemia initiating stem cells (LICs). It will also examine the activaion of negative feedback pathways and will define the effects of combinations of dual mTORC2/mTORC1 agents with different modulators of feedback loops, to promote elimination of LICs. Altogether, these studies should advance our understanding of the mechanisms of BCR-ABL-mediated leukemogenesis and provide the rationale for future clinical-translational efforts involving the use of dual mTORC1/2 catalytic inhibitors and/or AMPK activators for the treatment of resistant CML and Ph+ ALL.